RESUMEN
The toxicity of glycol ethers is associated with their oxidation to the corresponding aldehyde and alkoxyacetic acid by cytosolic alcohol dehydrogenase (ADH; EC 1.1.1.1.) and aldehyde dehydrogenase (ALDH; 1.2.1.3). Dermal exposure to these compounds can result in localised or systemic toxicity including skin sensitisation and irritancy, reproductive, developmental and haemotological effects. It has previously been shown that skin has the capacity for local metabolism of applied chemicals. Therefore, there is a requirement to consider metabolism during dermal absorption of these compounds in risk assessment for humans. Cytosolic fractions were prepared from rat liver, and whole and dermatomed skin by differential centrifugation. Rat skin cytosolic fractions were also prepared following multiple dermal exposure to dexamethasone, ethanol or 2-butoxyethanol (2-BE). The rate of ethanol, 2-ethoxyethanol (2-EE), ethylene glycol, 2-phenoxyethanol (2-PE) and 2-BE conversion to alkoxyacetic acid by ADH/ALDH in these fractions was continuously monitored by UV spectrophotometry via the conversion of NAD+ to NADH at 340 nm. Rates of ADH oxidation by rat liver cytosol were greatest for ethanol followed by 2-EE >ethylene glycol >2-PE >2-BE. However, the order of metabolism changed to 2-BE >2-PE >ethylene glycol >2-EE >ethanol using whole and dermatomed rat skin cytosolic fractions, with approximately twice the specific activity in dermatomed skin cytosol relative to whole rat skin. This suggests that ADH and ALDH are localised in the epidermis that constitutes more of the protein in dermatomed skin than whole skin cytosol. Inhibition of ADH oxidation in rat liver cytosol by pyrazole was greatest for ethanol followed by 2-EE >ethylene glycol >2-PE >2-BE, but it only inhibited ethanol metabolism by 40% in skin cytosol. Disulfiram completely inhibited alcohol and glycol ether metabolism in the liver and skin cytosolic fractions. Although ADH1, ADH2 and ADH3 are expressed at the protein level in rat liver, only ADH1 and ADH2 are selectively inhibited by pyrazole and they constitute the predominant isoforms that metabolise short-chain alcohols in preference to intermediate chain-length alcohols. However, ADH1, ADH3 and ADH4 predominate in rat skin, demonstrate different sensitivities to pyrazole, and are responsible for metabolising glycol ethers. ALDH1 is the predominant isoform in rat liver and skin cytosolic fractions that is selectively inhibited by disulfiram and responds to the amount of aldehyde formed by the ADH isoforms expressed in these tissues. Thus, the different affinity of ADH and ALDH for alcohols and glycol ethers of different carbon-chain length may reflect the relative isoform expression in rat liver and skin. Following multiple topical exposure, ethanol metabolism increased the most following ethanol treatment, and 2-BE metabolism increased the most following 2-BE treatment. Ethanol and 2-BE may induce specific ADH and ALDH isoforms that preferentially metabolise short-chain alcohols (i.e. ADH1, ALDH1) and longer chain alcohols (i.e. ADH3, ADH4, ALDH1), respectively. Treatment with a general inducing agent such as dexamethasone enhanced ethanol and 2-BE metabolism suggesting induction of multiple ADH isoforms.
Asunto(s)
Etanol/metabolismo , Glicoles de Etileno/metabolismo , Piel/metabolismo , Solventes/metabolismo , Alcohol Deshidrogenasa/antagonistas & inhibidores , Alcohol Deshidrogenasa/metabolismo , Disuasivos de Alcohol/farmacología , Aldehído Deshidrogenasa/antagonistas & inhibidores , Aldehído Deshidrogenasa/metabolismo , Animales , Citosol/enzimología , Citosol/metabolismo , Disulfiram/farmacología , Inhibidores Enzimáticos/farmacología , Etanol/química , Etanol/farmacocinética , Glicoles de Etileno/química , Glicoles de Etileno/farmacocinética , Técnicas In Vitro , Hígado/enzimología , Hígado/metabolismo , Masculino , NAD/metabolismo , Oxidación-Reducción/efectos de los fármacos , Pirazoles/farmacología , Ratas , Ratas Wistar , Piel/enzimología , Absorción Cutánea , Solventes/química , Solventes/farmacocinéticaRESUMEN
2-Butoxyethanol (2-BE) is widely used as an industrial solvent, which may result in human dermal exposure within the workplace. This study compares in vivo and in vitro skin absorption of 2-BE using similar application regimes and determines the potential of skin to metabolise this chemical prior to entering the systemic blood circulation. Following topical application of undiluted [1-14C] 2-BE to occluded rat skin in vivo, 28% of the dose was absorbed after 24 h. The major routes of excretion included the urine (19%), expiration as carbon dioxide (6%) and faeces (0.4%) whilst little of the dose remained in the carcass (1.3%). Free 2-BE (0.5%), butoxyacetic acid (8%), glucuronide conjugate (3%), sulphate conjugates (0.7%) and ethylene glycol (0.6%) were detected in urine. Permeation rates of 2-BE through unoccluded rat dermatomed skin (16%) were greater than rat whole skin (8%) whilst absorption through human dermatomed skin (4%) was lower than the rat. Absorption of undiluted 2-BE through occluded rat dermatomed skin in vitro (18%) most accurately predicted absorption through rat skin in vivo. However, 2-BE absorption (23%) was enhanced by application in methanol. Distribution analysis and microautoradiography demonstrated the lack of 2-BE accumulation within the skin in vitro or in vivo. This was reflected in the absence of first pass metabolism of 2-BE during percutaneous penetration through viable human or rat skin in vitro or rat skin in vivo, despite rat skin cytosol having the potential to metabolise 2-BE. In conclusion, the in vitro system provided a reasonable estimate of dermal absorption in vivo for the rat. Therefore, by extrapolation of the comparative in vitro data for human and rat skin in vitro, dermal absorption of 2-BE in man was about one-fifth of that in the rat. However, the rapid penetration through skin in vitro prevented local metabolism and systemic exposure after skin contact with 2-BE in vivo was likely to be to the parent compound. Thus, in vitro skin systems can be used to model dermal absorption of volatile glycol ethers, to predict how much compound enters the circulation and allows the toxicologist to evaluate the body burden of a chemical and potential systemic toxicity.
Asunto(s)
Glicoles de Etileno/metabolismo , Absorción Cutánea/fisiología , Piel/metabolismo , Administración Cutánea , Animales , Radioisótopos de Carbono , Glicoles de Etileno/farmacocinética , Glicoles de Etileno/orina , Heces/química , Humanos , Técnicas In Vitro , Inactivación Metabólica , Masculino , Ratas , Ratas WistarRESUMEN
Percutaneous absorption and cutaneous metabolism of 2-ethoxyethanol were assessed in vivo and with an in vitro flow-through diffusion system. Topical application of undiluted (14)C-ethoxyethanol to occluded rat skin in vivo resulted in 25% of the dose being absorbed after 24 h. The major routes of excretion included the urine (15%), expiration as carbon dioxide (6%), and feces (1.2%), while little of the dose remained in the carcass (1.3%). Free ethoxyethanol, ethoxyacetic acid, and glycine conjugate were detected in urine. Permeation rates of ethoxyethanol through unoccluded rat split skin (20%) were greater than rat whole skin (11%), while absorption through human split skin (8%) was lower than the rat. Absorption of undiluted ethoxyethanol through occluded rat split skin in vitro (22%) most accurately predicted absorption through rat skin in vivo. However, ethoxyethanol absorption (29%) was enhanced by application in methanol. First pass metabolism of ethoxyethanol was not detected during percutaneous penetration through viable human or rat skin in vitro or rat skin in vivo. However, rat skin cytosol had the potential to metabolize ethoxyethanol, suggesting that the rapid penetration through skin in vivo prevented metabolism and that systemic exposure after skin contact with 2-ethoxyethanol is likely to be to the parent compound. In conclusion, the in vitro system provided a reasonable estimate of dermal absorption for the rat in vivo and comparison of human and rat skin in vitro indicated 2-ethoxyethanol absorption in humans is about one-third of that in the rat.